title: Engineered short branched-chain acyl-CoA synthesis in E. coli and acylation of chloramphenicol to branched-chain derivatives.
abstract: Short branched-chain acyl-CoAs are important building blocks for a wide variety of pharmaceutically valuable natural products. Escherichia coli has been used as a heterologous host for the production of a variety of natural compounds for many years. In the current study, we engineered synthesis of isobutyryl-CoA and isovaleryl-CoA from glucose in E. coli by integration of the branched-chain α-keto acid dehydrogenase complex from Streptomyces avermitilis. In the presence of the chloramphenicol acetyltransferase (cat) gene, chloramphenicol was converted to both chloramphenicol-3-isobutyrate and chloramphenicol-3-isovalerate by the recombinant E. coli strains, which suggested successful synthesis of isobutyryl-CoA and isovaleryl-CoA. Furthermore, we improved the α-keto acid precursor supply by overexpressing the alsS gene from Bacillus subtilis and the ilvC and ilvD genes from E. coli and thus enhanced the synthesis of short branched-chain acyl-CoAs. By feeding 25 mg/L chloramphenicol, 2.96 ± 0.06 mg/L chloramphenicol-3-isobutyrate and 3.94 ± 0.06 mg/L chloramphenicol-3-isovalerate were generated by the engineered E. coli strain, which indicated efficient biosynthesis of short branched-chain acyl-CoAs. HPLC analysis showed that the most efficient E. coli strain produced 80.77 ± 3.83 nmol/g wet weight isovaleryl-CoA. To our knowledge, this is the first report of production of short branched-chain acyl-CoAs in E. coli and opens a way to biosynthesize various valuable natural compounds based on these special building blocks from renewable carbon sources.
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 Xiqidao, Airport Economic Park, Tianjin, 300308, China.
authors.forename: Huiping
authors.initials: H
authors.lastname: Bi
authors.forename: Yanfen
authors.initials: Y
authors.lastname: Bai
authors.forename: Tao
authors.initials: T
authors.lastname: Cai
authors.forename: Yibin
authors.initials: Y
authors.lastname: Zhuang
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Liang
authors.forename: Xueli
authors.initials: X
authors.lastname: Zhang
authors.forename: Tao
authors.initials: T
authors.lastname: Liu
authors.forename: Yanhe
authors.initials: Y
authors.lastname: Ma
mesh_terms: D000214:Acyl Coenzyme A / Q000378:metabolism*; D000215:Acylation; D000900:Anti-Bacterial Agents / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D001711:Biotransformation; D002701:Chloramphenicol / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics
chemical_list: D000214:Acyl Coenzyme A; D000900:Anti-Bacterial Agents; D011994:Recombinant Proteins; D002701:Chloramphenicol